Facilitated recycling pathway for RNA polymerase III

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• RNA polymerase III (pol III) is crucial for transcribing small RNAs.
• Understanding the mechanisms behind pol III's high in vitro transcription efficiency is key.

Purpose of the Study:

• To elucidate the primary mechanism driving the high in vitro transcription efficiency of yeast RNA polymerase III.
• To investigate the kinetics and requirements for RNA polymerase III recycling and reinitiation.

Main Methods:

• Kinetic analysis of RNA polymerase III transcription.
• Template competition assays.
• Heparin resistance assays to assess polymerase release.

Main Results:

• RNA polymerase III recycling on preassembled transcription complexes is significantly faster than the initial transcription cycle.
• Efficient recycling requires termination at the natural signal and is favored by high UTP concentrations; runoff transcription hinders recycling.
• Reinitiation exhibits increased resistance to heparin, indicating polymerase may not fully release after termination.
• Template competition assays demonstrate RNA polymerase III commitment to reinitiating on the same gene.

Conclusions:

• The high in vitro efficiency of yeast RNA polymerase III is primarily attributed to rapid polymerase recycling.
• A model proposing direct transfer of RNA polymerase from termination to promoter sites is supported by the findings.